The administration of dibutyryl cAMP (dbcAMP) causes some animal tumors to cease growing, suggesting that cAMP inhibits tumor growth. It has been suggested that in those tumors which are not inhibited by dbcAMP, the resistance is due to a deficiency in intracellular binding of cAMP. Consistent with this hypothesis, a dbcAMP-resistant variant of the Morris hepatoma has been found to bind less cAMP than do dbcAMPinhibitable variants. We have found that in normal rat liver cytosol and nuclei, GTP, in physiologic concentration (10-4 M), stimulates specific binding of [3H] cAMP to an Mr 57,000 cAMP binding protein (BP), which is presumably the regulatory subunit of protein kinase (PK) II. The effect of GTP requires a trypsin-labile cofactor, which can be removed using GTP agarose,a nd can be recovered from the agarose. In a preliminary study, we have observed a decrease in the effect of 10-4 M GTP on cAMP binding in a nuclear extract from a dbcAMP-resistant hepatoma, as compared to a dbcAMP responsive one. In the proposed project, we wish to explore the hypothesis that in hepatomas that are not inhibited by dbcAMP, the resistance is due to a decrease in the binding of cAMP to the GTP-stimulatable cAMP BP. Studies will be done to elucidate further the mechanism by which GTP enhances binding of cAMP in normal rat liver. We will attempt to purify the GTP cofactor by gel chromatography, and characterize it by studying its ability to bind [3H] guanosine imidodiphosphate ([3H] GIDP); to hydrolyze GTP; to enhance cAMP binding to purified PK II receptor in the presence of GTP; and to increase the PK activity of the purified PK II complex. We will also study GTP stimulation of cAMP binding in nuclear extracts and cytosol from rat hepatocytes maintained in primary culture,and from several rat hepatoma variants maintained in tissue culture. The effect of dbcAMP on the proliferation of these hepatomas will also be studied. If GTP-stimulatable cAMP binding is lower in the dbcAMP-resistant tumors than in the responsive one, or if it is lower in the tumors than in the hepatocytes, we will investigate whether this can be explaind by a decrease in the amount of the GTP-stimulatable BP or its affinity for cAMP; a decrease in the intrinsic responsiveness of the BP to GTP and added cofactor; or a change in the properties of the cofactor in these tumors.